Malaria continues to be a major world health problem, killing1-2 million people annually. The Plasmodium parasites that cause malaria have a multistage life cycle, which contributes to their ability to evade host immune responses. For malaria transmission, the parasite must undergo sexual differentiation into mature gametocytes that, when taken up in a blood meal by a mosquito can fertilize and begin sporogonic development. The molecular basis for the critical switch from asexual replication to sexual differentiation in the parasite's life cycle is unknown. Our hypothesis is that this transition is initiated by the activation of transcription factors that regulate the expression of genes required to effect gametocytogenesis. Once the regulatory factors are identified, inhibitors could be designed to block gametocytogenesis, which would prevent malaria transmission. To identify the genes involved, Plasmodium falciparum gametocyte producing (G+) and non-producing (G-) parasite lines were derived from strain 3D7 parasites. The first Specific Aim will be to use whole genome microarray analysis to compare mRNA harvested from the G+ and G- clones during the asexual to sexual transition. This comparison should identify the earliest differentially expressed genes. Specific Aim 2 will be to analyze the factors involved in regulating the gametocyte-specific expression of these genes. The ability of the 5'and 3'flanking regions of the genes to drive stage-specific reporter gene expression will be tested and the specific regulatory regions required will be identified by promoter mapping supplemented with motif analysis. Specific Aim 3 will be to identify the genes required for the induction of gametocytogenesis by determining the genetic differences between the G+ and G- parasite lines, as well as the genes that interact with the regulatory regions identified in Specific Aim 2. The relationship between gametocytogenesis and the genetic differences found will be further analyzed by complementation of the G- line and targeted gene disruption of wild type G+ parasites. This information should provide new gene candidates that could be used in the future to design new strategies to control malaria transmission.